Project Summary: Coenzyme Q is required for cellular aerobic function in all organisms, and is an important lipid soluble antioxidant. Several pathological states in humans, including cancer and changes associated with aging, implicate aberrant coenzyme Q metabolism or synthesis. Among the genes required for coenzyme Q production in yeast, coq-3 is required in two O-methyltransferase steps. In the roundworm Caenorhabditis elegans, strains with mutated coq-3 genes demonstrate slow development, sterility and poor viability. However, supplementing worm media with 4-methyl sterols such as lophenol may be able to rescue these coq-3 deficient worms. Nematodes are unable to synthesize sterols de novo, and rely on dietary cholesterol for their sterol needs. Among the potential modifications worms make to cholesterol, methylation of the position 4 carbon is an alteration unique to nematodes. Since coq-3 worms lack a gene with putative coenzyme Q methylation function, the partial rescue of these mutants by supplementation of a 4-methyl sterol implies that coq-3 in worms may function in both an O-methyltransferase and C- methyltransferase roles, either in complex with other polypeptides or alone. The proposal presented here seeks to elucidate the function of C. elegans COQ3 by evaluating its coenzyme Q O-methyltransferase activity, using a yeast strain bearing the worm coq-3 gene. Sterol carbon 4 C-methyltransferase activity will also be assessed, in yeast and worm mitochondria. Sterol and coenzyme Q concentration will be measured via HPLC, and GC-MS will be used to identify sterol products and concentrations. Rescue of coq-3 mutant worms will be assayed by growth in media supplemented with either lophenol or cholesterol. Brood sizes and lifespans will be used to indicate the presence and degree of rescue. Aerobic activity will also be used to evaluate rescue. Since methylation of the carbon 4 position in sterols is unique to nematodes, discovery of the candidate enzyme responsible may serve as a potent nematocidal drug. Relevance: In humans, coenzyme Q and cholesterol share a common biosynthetic pathway. Roundworms may involve these pathways in a different manner than people, and these differences can provide scientists with targets for drugs against parasitic and agriculturally destructive roundworms.